The invention relates to protein kinases and methods of activating or inhibiting the expression of protein kinases.
Mitogen-activated protein kinase (MAPK) cascades have been remarkably conserved in evolution. The core of these cascades is a three-tiered module of serine/threonine kinases that consists of a MAPK-extracellular signal regulated kinase kinase (a MEKK), a MEK, and a MAPK or extracellular signal regulated kinase (ERK). In simple eukaryotes, such as the budding yeast Saccharomyces cerevisiae (S. cerevisiae), and the fission yeast, Saccharomyces pombe (S. pombe), these cascades are activated predominantly by cellular stresses such as nutritional starvation and osmolar stress (reviewed in Elion, TIBS 5:322 (1995); Herskowitz, Curr. Oper. Cell 80:187 (1995); and Levin et al., Cell Biol. 7:197 (1995)). In mammals, these cascades have evolved to allow responses to complex stimuli (e.g., growth factors and inflammatory cytokines), but in many cases, such as the response to osmolar challenge (Galcheva-Gargova et al., Science 265:806 (1994); Han et al., Science 265:808 (1994)), the primitive stress responses remain intact.
Epistasis analyses in yeast suggest that the Sterile 20 (Ste20) protein serine/threonine kinases and related protein kinases act upstream of the three tiered module. Three mammalian homologs of Ste20 have been reported to date: p21-activated protein kinase (PAK1) and related PAKs (Manser et al., Nature 367:40 (1994); Martin et al., EMBO J. 14:1970 (1995)); germinal center (GC) kinase (Katz et al., J. Biol. Chem. 269:16802 (1994)); and mammalian Ste20-like kinase 1 (MST1) (Creasy et al., J. Biol. Chem. 270:21695 (1995)). Mammalian Ste20s may function upstream of MEKK/MEK/MAP kinase pathways. PAK1 (Manser et al., Nature 367: 40 (1994)) and GC kinase (Katz et al., J. Biol. Chem. 269:16802 (1994)) have been shown to be capable of activating mammalian MAPK kinases (Polverino et al., J. Biol. Chem. 270:26067 (1995); Pombo et al., Nature 377:750 (1995); Zhou et al., J. Biol. Chem. 270:12665 (1995)), further illustrating remarkable evolutionary conservation of the MAPK kinases. When co-transfected with MAP kinase, both PAK1 and GC kinase activate the stress-activated protein kinase (SAPK)/c-Jun amino terminal kinase (JNK) cascade. PAK1 also activates the stress activated MAPK, p38, as well.
Ste20 protein kinases can be divided into two families based on their structure and regulation. The first family is the Ste20 family, which includes Ste20, PAK1 and related PAKs. These proteins contain a carboxy terminal catalytic domain and an amino terminal regulatory domain which has a p21.sup.cdc42/rac1 binding region (Manser et al., Nature 367:40 (1994); Martin et al., EMBO J. 14:1970 (1995)). PAK1 appears to be activated by binding to cdc42Hs or Rac1. Following binding to the small GTP-binding proteins, the kinase undergoes autophosphorylation and is activated. Physiologic activators of PAK1 have been identified, and include the chemoattractant peptide fMetLeuPhe, and Interleukin 1 (IL-1) (Zhou et al., J. Biol. Chem. 270:12665 (1995)).
The second family of Ste20s is the Sps1 family. Members of this group include Sps1, which is encoded by the S. cerevisiae sporulation specific 1 gene, which is necessary for spore formation in response to nutritional starvation; and the mammalian genes MST1 and GC kinase. The catalytic domain is amino terminal in these proteins, and the function of their carboxy terminal regions has not previously been known. These kinases do not contain an identifiable Rac/cdc42Hs binding domain in their non-catalytic regions. The regulation of this family of Ste20s is not well characterized. MST1 appears to be activated by dephosphorylation. Sps1 and its MAPK, Smk1 (Krisak et al., Genes & Development 8:2151 (1994)), are transcriptionally regulated, being expressed only at certain stages of the sporulation process, but it is not known if there are other modes of regulation of Sps1. Physiological activators of the Sps1 family of Ste20s have not been previously identified.